This project is concerned with the regulation of intracellular activities of Ca, Na and H in mammalian myocardium, the sources of Ca which support contraction and the mechanisms by which this Ca is delivered to the myofilaments. Using ion-selective microelectrodes (ISMEs) one can monitor directly the intracellular activities of the ions of interest (e.g., Ca, Na and H). These results will be correlated with biochemical and flux studies to examine more comprehensively the parameters concerned. One hypothesis to be tested is whether the model that the inotropic response of cardiac glycosides is due to a Na-pump inhibition and subsequent increase in aNa(i) and aCa(i) (via Nai-Ca(o) exchange). Are there discrepancies at very low glycoside concentrations or with respect to time and is diastolic aCa(i) increased in inotropic conditions. Another related area then is determining the characteristics of the Na-Ca exchange system in the intact cell to discern whether these could be consistent with this system playing a significant role in Ca influx during a beat. Experiments are also planned to investigate the extent and importance of intracellular compartmentalization of Ca and Na. Another area is involved with elucidation of which sources of Ca (SR, sarcolemmal, mitochondrial) support tension development. It is hypothesized that there may be significant variation in which sources of contractile dependent Ca predominate in different cardiac preparations. The study of different cardiac preparations with respect to [Ca]o, frequency and pharmacological interventions may provide further insight into the relative importance of SR and sarcolemmal Ca in tension development in different cardiac tissues. The potential role of mitochondria in control of steady state aCa(i) and aNa(i) and compartmentalization of Na and Ca will bw studied. The relationship between aCa(i) and tension will be examined. The effects of certain agents (e.g., catecholamines, methylaxanthines, verapamil) on aCai will also be evaluaated. Also the interrelations between PH(i) and aCa(i) during anoxia, ischemia and recovery will be investigated. The studies in this project should thus increase our understanding of basic myocardial function in normal and pathologic conditions.